Differential gene expression plays a critical role in the development of multicellular organisms. At a 'systems level' (e.g. at the level of an organ, tissue or whole organism), this process can be studied using gene regulatory network (GRN) models that capture physical and regulatory interactions between genes and their regulators, e.g. transcription factors (TFs), transcriptional cofactors and microRNAs. In the past years, significant progress has been made toward the characterization of developmental GRNs. However little is known about the organization and function of GRNs that control post-developmental physiology. Differential gene expression likely plays an important and broad role in different aspects of organismal physiology (e.g. homeostasis, response to environmental cues, hereafter referred to as 'systems physiology'). This is because the majority of TFs are expressed throughout the lifetime of an organism, which strongly suggests that they function in post-developmental processes. However, only few TFs involved in systems physiology have been identified and/or characterized. We use the digestive tract of the nematode C. elegans as a model system to study metazoan GRNs. The digestive tract is composed of the pharynx, intestine and rectum, its development has been well documented, and multiple regulators expressed in this system have already been identified. The digestive tract plays many important roles in C. elegans physiology; it is not only important for digestion and metabolism, it is also a major interface between the animal and the environment. For instance, it responds to different environmental stressors and provides the innate immune response upon exposure to pathogens. Thus, the digestive tract is an ideal model to study GRNs that pertain to systems physiology. In the proposed project, we will focus on the following questions: 1) Which regulators of differential gene expression are expressed in the C. elegans digestive tract? 2) How are these regulators regulated? 3) How is the expression of genes involved in physiological functions, such as metabolism, stress response and innate immunity controlled? And 4) What is the relative contribution of different 'genic' regions (e.g. promoters, exons, introns and enhancers) to differential gene expression in the digestive tract? We, and others, have generated many C. elegans resources and tools that uniquely enable us to carry out the proposed project. These include high-quality TF predictions, high-throughput gene-centered yeast one-hybrid (Y1H) assays for the delineation of systems-level GRNs and two complementary full-length TF clone collections that can be used in high-throughput Y1H assays and RNAi experiments, respectively. PUBLIC HEALTH RELEVANCE: Differential gene expression plays a critical role in organism development, homeostasis and response to physiological or environmental cues, as well as in a multitude of diseases such as cancer, obesity and diabetes. At a systems level, differential gene expression can be studied using gene regulatory network (GRN) models that capture physical and regulatory interactions between genes and their regulators. We will continue our studies of digestive tract GRNs in the nematode C. elegans. Importantly, many genes and functions are highly conserved between worms and humans, and thus the C. elegans digestive tract provides a highly suitable model for metazoan GRN analysis. Our studies will provide insights into the functions of GRNs during development and in a wide variety of physiological functions such as stress response, metabolism and fighting pathogenic infection.